Imaging biomarkers of inflammation <Emphasis Type="Italic">in situ</Emphasis> with fun ctionalized quantum dots in the dextran sodium sulfate (DSS) model of mouse colitis

Objective and design: Myeloperoxidase (MPO) and proinflammatory cytokines play an important role in the development of inflammation. These markers are generally measured using tedious ELISA procedures. In this study, a novel technique utilizing antibody conjugated quantum dot nanoparticles was developed to detect Myeloperoxidase, Interleukin-1α (IL-1α) and Tumor Necrosis Factor-α (TNF-α) in vivo in the dextran sodium sulfate (DSS) model of experimental colitis. Materials and methods: Colitis was induced in animals (n = 8 animals/group) by feeding 4% DSS solution ad libitum for seven to eight days. Quantum Dots (QDs) exhibiting fluorescence at various wavelengths were conjugated to MPO, IL-1α and TNF-α polyclonal antibodies and tested in vivo at various stages of colitis. Tissue sections obtained were imaged with confocal microscope. The image intensity obtained from the tissue specimen was correlated with clinical activity measured as Disease Activity Index (DAI). Results: Myeloperoxidase, IL-1α and TNF-α were visualized with quantum dots on various days of disease. The intensity of quantum dots increased with the increase in inflammation. The increase in intensity showed an excellent correlation with the DAI based on the clinical parameters. Conclusion: The study demonstrated that multiple biomarkers can be detected simultaneously and their quantitative expression correlated well with clinical disease severity. This novel technology should facilitate design of a novel optical platform for imaging various biomarkers of inflammation, early detection of acute and chronic disease markers and inflammation-mediated cancer markers. This detection may also facilitate determination of therapeutic success. Received 14 March 2007; returned for revision 8 May 2007; accepted by M. Parnham 27 June 2007     

Local injection of a mixture of beta-aminopropionitril fumarate and sodium hyaluronate together with controlled exercise for treatment of subacute superficial digital flexor tendonitis in horses

Local injection of a mixture of beta-aminopropionitril fumarate (BAPN-f) and sodium hyaluronate (NaH⁺) together with controlled exercise were evaluated for treatment of superficial digital flexor tendon (SDFT) injuries in horses. Fourteen mixed breed horses with subacute SDF tendon injuries in forelimbs were randomly assigned to two groups. One group received BAPN-f (0.7 mg/ml) and the other received NaH⁺ (10 mg/ml) all by intratendinous injection. Controlled exercise started during the first week after intratendinous treatment and continued for 12 weeks. Cross-sectional area (CSA) of the SDFT, diameter of the SDFT, and relative area of the lesion (presence of CSA) were measured by ultrasonographic examination. Lesions were semiquantitatively graded for echogenicity on a scale of 0 to 4. The lesion severity in CSA was significantly reduced by BAPN and NaH compared to those horses treated by BAPN only (p < 0.05). Lesion echogenicity score was significantly higher in horses treated with BAPN and NaH⁺ at day 0 compared to horses treated with BAPN (p < 0.05). At the end of the study, lesion echogenicity score was significantly reduced in the horses treated with BAPN and NaH⁺ compared to horses treated by BAPN only (p < 0.05). According to the results of the present study, a combination of BAPN-f and NaH⁺ has a greater beneficial effect on tendon healing and remodeling in horses, as assessed by sonographic examination, compared to treatment with single drugs.     

急性心肌梗死后1周兔心室肌细胞钠离子通道活性的变化

目的:研究急性心肌梗死(AMI)后心室肌细胞钠离子通道活性的变化。方法:采用结扎兔冠状动脉左前降支方法建立AMI动物模型, 应用膜片钳全细胞记录方法, 观察AMI后1周心外膜梗死区心肌细胞钠通道电流(I_Na)的变化。结果:正常对照组I_Na电流密度峰值(-30mV)为(45.5±5.33)PA/PF(n=12), 心梗组为(22.48±4.62)PA/PF(n=14), 明显低于对照组, P＜0.01, I_Na电流-电压关系曲线在心梗组明显下移;心梗组I_Na失活曲线较对照组明显左移, 对照组V_0.5为(-76.2±5.3)mV(n=5), 心梗组为(-89.1±5.6)mV(n=6), P＜0.05;心梗组钠通道灭活后恢复时程明显慢于对照组, 恢复曲线下移。结论:AMI可导致梗死区心室肌细胞I_Na下降、钠通道动力学发生变化, 引起心肌传导速度下降和不应性延长, 可能是导致AMI后出现折返性室性心律失常的原因。     

Dual effects of melatonin on barbiturate-induced narcosis in rats

Melatonin affects the circadian sleep/wake cycle, but it is not clear whether it may influence drug-induced narcosis. Sodium thiopenthal was administered intraperitoneally into male rats pre-treated with melatonin (0.05, 0.5, 5 and 50 mg/kg). Melatonin pre-treatment affected in a dual manner barbiturate narcosis, however, no dose-effect correlation was found. In particular, low doses reduced the latency to and prolonged the duration of barbiturate narcosis. In contrast, the highest dose of melatonin (50 mg/kg) caused a paradoxical increase in the latency and produced a sustained reduction of the duration of narcosis, and a reduction in mortality rate. Melatonin 0.5 and 5 mg/kg influenced the duration but not the latency of ketamine- or diazepam-induced narcosis. Thus, the dual action of melatonin on pharmacological narcosis seems to be specific for the barbiturate mechanism of action.     

Effect of increased distal sodium delivery on organic osmolytes and cell electrolytes in the renal outer medulla

Sodium absorption in distal tubule segments was stimulated by increasing the distal delivery via infusion of hypertonic saline. In these animals, and in control rats, electrolyte concentrations in thick ascending limb cells, light and dark cells of the collecting duct in the outer and inner stripe of the outer medulla and in cells of the proximal straight tubule (outer stripe only) were studied. The measurements were performed by electron microprobe analysis of freeze-dried cryosections of the outer medulla. In addition, organic osmolytes (glycerophosphorylcholine, betaine and myo-inositol) were measured by high performance liquid chromatography in cortex and outer medulla. Augmented delivery of sodium chloride to the distal tubule was associated with increased sodium concentrations of thick ascending limb cells both in the outer and inner stripe and of medullary collecting duct light and dark cells in the outer stripe. While the sum of organic osmolyte concentrations was 28% higher in the outer medulla of the salt-loaded animals compared with controls, this value was unchanged in the renal cortex. These findings indicate that the primary event underlying stimulation of sodium absorption along the thick ascending limb during increased distal sodium delivery is enhanced entry of sodium across the apical cell membrane. This would be expected to lead to higher cell sodium concentrations and stimulation of basolateral active Na-K-exchange. The enhanced transport activity of outer medullary tubules may be associated with increased interstitial tonicities and intracellular retention of organic osmolytes.     

Hyperpolarizing current of the Na/K ATPase contributes to the membrane polarization of the Purkinje cell in rat cerebellum

 The contribution of the Na/K ATPase (pump) current to the polarization of the Purkinje cell has been studied using slices of the rat cerebellum by blocking the pump with dihydro-ouabain (DHO) while recording the membrane potential with microelectrodes in the somata. From our recordings, it appeared that blocking the pump depolarized the Purkinje cells more rapidly than might be expected from shifts in Na⁺ and K⁺ concentrations, suggesting the removal of a hyperpolarizing current. Application of DHO, in the presence of tetrodotoxin (TTX), led to calcium spike firing and plateau-like discharges suggesting activation of voltage-dependent calcium channels in the dendrites. Adding 2 mM Co²⁺ to the medium did not prevent the depolarizations. Removing calcium from the bathing medium containing 2 mM Co²⁺ blocked the spiking activity but DHO application still produced a depolarization. Experiments to measure the current inhibited by DHO indicated that the Na/K pump supplies a constant current of 240 pA. Substitution of the sodium with choline produced a hyperpolarization, during which DHO had no effect on the membrane potential. Substitution of the sodium with lithium produced only a slowly developing depolarization. It is concluded that in the cerebellar Purkinje cell, a continuous sodium ion influx activates the pumps which produce a current that directly contributes to the membrane polarization. Possible pathways for this sodium influx are discussed. Received: 3 April 1997 / Received after revision and accepted: 12 May 1997     

Potassium induced potential changes in rat diaphragm muscle

Summary The effect of different potassium concentrations on the membrane potential and membrane resistance of rat diaphragm muscle fibres was measured by means of a double sucrose gap method and a microelectrode technique. Concentration measurements showed that the muscle fibres gained sodium and lost potassium in the equilibration period.In the absence of external chloride changing the external potassium concentration from 2.8 mM to potassium-free caused a depolarization of the membrane of about 30 mV and a small increase in membrane resistance.This K-dependent potential change (K-response) was reduced by ouabain, K-strophanthin, 2,4-dinitrophenol and cyanide, indicating that an energy requiring process is involved. The temperature dependence of the K-response found is consistent with this assumption. Variation in potassium permeability in the absence and presence of external potassium could account for only 13% of the K-response.The K-response amplitude appeared to depend on the external potassium and the internal sodium concentration. Hyperpolarization of the membrane could not only be produced after readmission of potassium but also after addition of thallium, the latter being more potent. Raising the external chloride concentration resulted in a decrease of the K-response and membrane resistance. The current, generating the Kresponse was shown to be hardly influenced by conditional polarization of the membrane.It is concluded from these results that the Kresponse is mainly due to the operation of an electrogenic sodium pump.     

The effects of sodium butyrate,an inhibitor of histone deacetylase,on the cocaine- and sucrose-maintained self-administration in rats

In order to substantiate the concept that cocaine behavioral effects may be influenced by histone modification, rats were trained to self-administer cocaine intravenously (0.75 mg/(kg injection)), and were systemically pretreated with sodium butyrate (NaBu), a potent histone deacetylase inhibitor, before the test session during the maintenance phase. The effect of NaBu on a control reinforcer (sucrose)-induced self-administration was also assessed. NaBu (100–200 mg/kg) was inactive in altering the cocaine (0.75 mg/(kg injection))-maintained responding and at the highest dose (400 mg/kg) it did increase cocaine-induced lever presses during the maintenance phase. On the other hand, sucrose-reinforcing potential was not altered when NaBu was given at the highest dose (400 mg/kg). These findings extend previous observations that changes in histone acetylation are relevant to cocaine-induced behavioral effects. Given that histone acetylase inhibitor enhances cocaine-induced behavioral plasticity, the therapeutic benefits of histone acetyltransferase inhibitors warrant further investigation in the experimental models of cocaine abuse.     

Effect of hypothermia on renal sodium reabsorption

Summary The relationship between sodium reabsorption and oxygen consumption was studied in an isolated rabbit kidney preparation perfused with blood at 37, 28 and 19° C. When the temperature was lowered from 37° C to 28° C and to 19°C the rate of oxygen consumption and of the maximal P.A.H. excretion (Tm P.A.H.) decreased more than that of sodium reabsorption.TheQ ₁₀ for sodium reabsorption is about 1.8, while that for maximal P.A.H. excretion is 2.5. Some hypothesis on the possible mechanisms of the lowQ ₁₀ of the Na⁺ reabsorption are forwarded.Preliminary reports have been published [Boll. Soc. Ital. Biol. Sper.43, 1019–1023 (1966) and44, 1784–1787 (1967);45, 860–862 (1969) and45, 863–865 (1969)].     

Active Ca2+ reabsorption in the proximal tubule of the rat kidney

Summary Using the stop flow microperfusion technique with simultaneous capillary perfusion the rate of active Ca²⁺ reabsorption was evaluated by measuring the static head electrochemical potential difference as well as the permeability of the tubular wall for Ca²⁺ ions. Under control conditions the active Ca²⁺ transport was calculated to be 3.35×10^–13 mol/cm·s. It declined toward zero if the ambient Na⁺ was replaced by choline or lithium. Parallel experiments in the golden hamster showed that active Ca²⁺ transport, vanished completely if active Na⁺ transport was blocked by ouabain (1 mM). These data indicate that the active Ca²⁺ reabsorption from the proximal tubule depends on the active reabsorption of Na²⁺ presumably via a Na⁺–Ca²⁺ countertransport at the contraluminal cell membrane. The static head electrochemical potential difference of Ca²⁺ is the same in late and early proximal tubules. It is also not affected by the presence of acetazolamide (10^–4 M) by the absence of bicarbonate or glycodiazine buffer or by the absence or presence of phosphate (2 mM).